Method of monitoring biofouling in membrane separation systems

ABSTRACT

Methods for monitoring and controlling biofouling in membrane separation systems are provided. The present invention uses measurable amounts of fluorogenic agent(s) added to a feed stream to monitor and control the level of microbial growth during membrane separation and, thus, the purification of such feed stream during membrane separation.

CROSS-REFERENCE TO RELATED PATENT APPLICATION

[0001] This patent application is a continuation-in-part of U.S. patentapplication Ser. No. 10/201,088, filed Jul. 23, 2002, now pending.

FIELD OF THE INVENTION

[0002] The present invention relates generally to membranes and, moreparticularly, to methods for monitoring and controlling biofouling inmembrane separation systems.

BACKGROUND OF THE INVENTION

[0003] Membrane separation, which uses a selective membrane, is a fairlyrecent addition to the industrial separation technology for processingof liquid streams, such as water purification. In membrane separation,constituents of the influent typically pass through the membrane as aresult of a driving force(s) in the feed stream, to form the permeatestream (on the other side of the membrane), thus leaving behind someportion of the original constituents in a stream known as theconcentrate.

[0004] Membrane separations commonly used for water purification orother liquid processing include microfiltration (MF), ultrafiltration(UF), nanofiltration (NF), reverse osmosis (RO), electrodialysis,electrodeionization, pervaporation, membrane extraction, membranedistillation, membrane stripping, membrane aeration, and otherprocesses.

[0005] Pressure-driven membrane filtration uses pressure as the drivingforce. Pressure-driven membrane filtration is also known as membranefiltration. Pressure-driven membrane filtration includesmicrofiltration, ultrafiltration, nanofiltration and reverse osmosis. Incontrast to pressure driven membrane filtration an electrical drivingforce is used in electrodialysis and electrodeionization.

[0006] Historically, membrane separation processes or systems were notconsidered cost effective for water treatment due to the adverse impactsthat membrane scaling, membrane fouling, membrane degradation and thelike had on the efficiency of removing solutes from aqueous waterstreams. However, advancements in technology have now made membraneseparation a more commercially viable technology for treating aqueousfeed streams suitable for use in industrial processes.

[0007] Furthermore, membrane separation processes have also been mademore practical for industrial use, particularly for raw and wastewaterpurification. This has been achieved through the use of improveddiagnostic tools or techniques for evaluating and monitoring membraneseparation performance. The performance of membrane separation, such asefficiency, for example: flux, membrane permeability, permeate recovery,energy efficiency, time between membrane cleanings or time to conduct acleaning cycle, and effectiveness, for example: rejection orselectivity, are typically reduced by membrane fouling.

[0008] Membrane separation processes are prone to fouling by microbes,which is known as biofouling. The growth of microorganisms duringmembrane separation is a constant concern particularly in aqueousstreams which provide optimum conditions for microbial growth.Biofouling is particularly detrimental to membrane separation systemsbecause once it is started, the growth rate accelerates and biofoulingcan facilitate other types of fouling as well. For example, theexopolymeric substances (“EPS”) or slime layer of the biomass can trapand hold scales and other particulates that might otherwise pass out ofthe membrane separation system during operation. Furthermore, a thickEPS layer can also decrease turbulent flow within the membrane. This canlead to an increase in the concentration polarization layer which is aknown contributor to membrane scaling phenomena.

[0009] The immediate and most obvious effect of biofouling is a decreasein membrane permeate output and/or a rise in the pressure drop along thelength of a membrane element on the feed and concentrate side of themembrane, referred to herein as “differential pressure.” Under aconstant pressure, this results in a loss in the production of permeate.Pressure can be increased in order to maintain a constant flux, but thisincreases energy consumption and further accelerates fouling. Inaddition, continued operation under these conditions (i.e., a loss ofpermeate flux, an increase in pressure differential and an increase inthe pressure driving force) would necessarily require an increasednumber of cleanings over the life time of the membrane, therebydecreasing the membrane life and potentially increasing water costs if asignificant amount of down time is required due to the cleanings. Lessobvious effects include reduced solute rejection, contamination ofpermeate and deterioration of membrane modules, such as biodegradationon membrane glue lines. The review article written by H. F. Ridgway & H.Flemming entitled “Membrane Biofouling”, Water Treatment MembraneProcesses, McGraw Hill, pp. 6.1 to 6.62, 1996, is helpful to anunderstanding of these matters.

[0010] In general, biofouling is controlled through the use of biocidesand other biocontrol agents, i.e., chemicals that can inhibit microbialgrowth by destroying the cell wall or cellular constituents ofmicroorganisms. Mechanical means and radiation means are additionalpossibilities. Intermittent use of biocides is typically encouragedsince biocides can be both expensive and toxic. Thus, to prevent waste,constant monitoring and testing of the water system and of the membraneprocess parameters are required to determine the proper quantity ofbiocide for controlling microbial growth.

[0011] However, known monitoring techniques may not provide an adequatelevel of sensitivity, specificity and/or accuracy with respect tomonitoring the effects of biofouling on membrane separation. Typicalmonitoring techniques include pressure and flow measurements and grabsampling to determine microbial population. With respect to pressuremeasurements, monitoring is generally conducted by evaluating changes inthe differential pressure along the length of the membrane. With respectto flow measurement, the flow meters generally employed in such systemsare subject to calibration inaccuracies, thus requiring frequentcalibration. However, the changes in pressure and flow are notnecessarily specific to biofouling, as they can be influenced by anysuitable increase in scalants, foulants and/or like constituents thatcan build-up and remain in the system during membrane separation. Aspreviously discussed, the microbial growth layer can enhance other typesof fouling since it can trap or hold scales and other particulates thatmight otherwise pass out of the system during membrane separation.

[0012] For grab samples, water samples are typically taken from the feedstream and the exit stream. Samples from the permeate stream can also betaken to determine if there is any contamination in the permeate. Atypical technique involves withdrawing a sample, diluting the sample,and applying the sample to the surface of a nutrient agar medium. Afterincubation for 24 to 28 hours, the sample is checked for the presence ofmicroorganisms and, where appropriate, the organisms are counted bymanual or video means. A variation on this method includes withdrawing asample and culturing it for a predetermined time, and then observing theculture medium by nephelometry or turbidimetry. In other words, thepresence of microorganisms is revealed by the opacity of the culturemedium.

[0013] A significant problem associated with grab sampling is the timelag between withdrawing the sample and completing the analysis todetermine the level of microbiological activity in the sample. In thisregard, the time lag can be exacerbated when the samples have to betransported off-site for analysis which can further delay obtaining theresults.

[0014] Another problem associated with grab sampling is that it canunderestimate the overall microbiological activity in the industrialwater system because grab sampling is only sufficient to provide anindication of the planktonic microbiological activity, not the sessileactivity. Planktonic microbiological populations are alive and existsuspended within the water of a water system. As used herein, the term“sessile” refers to populations of microorganisms that are alive, butimmobile. It is possible to get an industry-acceptable measurement ofplanktonic populations by grab sampling since planktonic microorganismsare suspended within the water sample that is removed and tested formicroorganism concentrations. In contrast, sessile populations arestrongly attached to the structures within the system and their presenceis not easily measured by removing a sample of water and testing thissample for microorganisms. In this regard, the level of planktonic cellsmay not directly correlate to the level of sessile cells in the membraneseparation system.

[0015] Accordingly, a need exists to monitor and/or control biofoulingin membrane separation systems in real-time where conventionalmonitoring techniques are generally complex and/or may lack thesensitivity, specificity and/or accuracy necessary to adequately monitorbiofouling such that membrane separation performance can be optimized.

SUMMARY OF THE INVENTION

[0016] The first aspect of the instant claimed invention is a method ofmonitoring biofouling in a membrane separation system including amembrane capable of separating a feed stream into at least a firststream, known as the permeate, and a second stream, known as theconcentrate, comprising the steps of:

[0017] (a) selecting a fluorogenic agent wherein the selection is madesuch that it is known in advance whether said fluorogenic agent is

[0018] (i) capable of traveling through the membrane into the permeatestream, or

[0019] (ii) not capable of passing through the membrane into thepermeate stream;

[0020] (b) adding the fluorogenic agent to the feed stream;

[0021] (c) providing one or more fluorometers to detect the fluorescentsignal of the fluorogenic agent in at least one of the feed stream, theconcentrate and optionally the permeate;

[0022] (d) allowing the fluorogenic agent to react with at least onemicroorganism within the membrane separation system to form a reactedfluorogenic agent;

[0023] (e) using said one or more fluorometers to detect the fluorescentsignal of at least one of the fluorogenic agent and the reactedfluorogenic agent in at least one of the feed stream and the concentrateand optionally the permeate; and

[0024] (f) using the fluorescent signal of at least one of thefluorogenic agent and the reacted fluorogenic agent to monitorbiofouling in the membrane separation system based on the change in thefluorescent signal of the fluorogenic agent, or the reacted fluorogenicagent or a combination of both fluorescent signals.

[0025] The second aspect of the instant claimed invention is a method ofmonitoring biofouling in a membrane separation system including amembrane capable of separating a feed stream into at least a firststream, known as the permeate, and a second stream, known as theconcentrate, comprising the steps of:

[0026] (a) selecting a fluorogenic agent wherein the selection is madesuch that it is known in advance whether said fluorogenic agent is

[0027] (i) capable of traveling through the membrane into the permeatestream, or

[0028] (ii) not capable of passing through the membrane into thepermeate stream;

[0029] (b) selecting an inert fluorescent tracer wherein the selectionis made such that it is known in advance whether said inert fluorescenttracer is

[0030] (i) capable of traveling through the membrane into the permeatestream, or

[0031] (ii) not capable of passing through the membrane into thepermeate stream;

[0032] (b) adding the fluorogenic agent and the inert fluorescent tracerto the feed stream, wherein said fluorogenic agent and said inertfluorescent tracer are added in a known proportion to each other;

[0033] (c) providing one or more fluorometers to detect the fluorescentsignal of the fluorogenic agent and the fluorescent signal of the inertfluorescent tracer in at least one of the feed stream or the concentrateor optionally the permeate;

[0034] (d) allowing the fluorogenic agent to react with at least onemicroorganism within the membrane separation system to form a reactedfluorogenic agent;

[0035] (e) using said one or more fluorometers to detect the fluorescentsignal of at least one of the fluorogenic agent and the reactedfluorogenic agent and the inert fluorescent tracer in at least one ofthe feed stream and the concentrate and optionally the permeate; and

[0036] (f) using the fluorescent signal of at least one of thefluorogenic agent and the reacted fluorogenic agent and the inertfluorescent tracer to monitor biofouling in the membrane separationsystem based on the change in the fluorescent signal of the fluorogenicagent, or the reacted fluorogenic agent or a combination of bothfluorescent signals relative to the fluorescent signal of the inertfluorescent tracer.

DETAILED DESCRIPTION OF THE PRESENTLY PREFERRED EMBODIMENTS

[0037] Throughout this patent application, the following terms have theindicated meanings.

[0038] “capable” means “having the ability or qualities necessary for acertain task”.

[0039] “pore” A pore is a pathway, which functions as the means by which“something” is intended to pass through a membrane surface. The“something” may be a solute, a particle, a molecule or anything of suchdimension that it is capable of fitting through the pore. A “pore” isdifferent than a “hole” in the membrane because “holes” or “defects” inthe membrane are not intended to be present in the membrane. A “pore”may be an opening of a defined size and shape, for example a cylindricalpore with a 0.2 micron diameter. Alternatively, a “pore” may be atorturous path consisting of a series of openings of undefined shapewhich allow particles of only a certain overall size to pass. The term“pore” is generally not in common usage with reverse osmosis membranesbecause with reverse osmosis membranes the “pores” may only be as smallas the interstitial voids between the polymer nodules in a polymermembrane.

[0040] “permeate” The permeate stream is that stream that travelsthrough the pores of the membrane.

[0041] “concentrate” The concentrate stream is that stream that does nottravel through the pores of the membrane.

[0042] “Nalco” refers to Nalco Company, 1601 W. Diehl Road, Naperville,Ill. 60563, (630) 305-1000.

[0043] The first aspect of the instant claimed invention is a method ofmonitoring biofouling in a membrane separation system including amembrane capable of separating a feed stream into at least a firststream, known as the permeate, and a second stream, known as theconcentrate, comprising the steps of:

[0044] (a) selecting a fluorogenic agent wherein the selection is madesuch that it is known in advance whether said fluorogenic agent is

[0045] (i) capable of traveling through the membrane into the permeatestream, or

[0046] (ii) not capable of passing through the membrane into thepermeate stream;

[0047] (b) adding the fluorogenic agent to the feed stream;

[0048] (c) providing one or more fluorometers to detect the fluorescentsignal of the fluorogenic agent in at least one of the feed stream, theconcentrate and optionally the permeate;

[0049] (d) allowing the fluorogenic agent to react with at least onemicroorganism within the membrane separation system to form a reactedfluorogenic agent;

[0050] (e) using said one or more fluorometers to detect the fluorescentsignal of at least one of the fluorogenic agent and the reactedfluorogenic agent in at least one of the feed stream and the concentrateand optionally the permeate; and

[0051] (f) using the fluorescent signal of at least one of thefluorogenic agent and the reacted fluorogenic agent to monitorbiofouling in the membrane separation system based on the change in thefluorescent signal of the fluorogenic agent, or the reacted fluorogenicagent or a combination of both fluorescent signals.

[0052] Membranes suitable for use in the instant claimed inventioninclude cross-flow systems, dead-end flow systems, reverse osmosis,nanofiltration, ultrafiltration, microfiltration, electrodialysis,electrodeionization, pervaporation, membrane extraction, membranedistillation, membrane stripping, membrane aeration and the like orcombinations thereof. The preferred membrane separation systems arereverse osmosis, nanofiltration, ultrafiltration and microfiltration

[0053] In reverse osmosis, the feed stream is typically processed undercross-flow conditions. In this regard, the feed stream flowssubstantially parallel to the membrane surface such that only a portionof the feed stream diffuses through the membrane as permeate. Thecross-flow rate is routinely high in order to provide a scouring actionthat lessens membrane surface fouling. This can also decreaseconcentration polarization effects (e.g., concentration of solutes inthe reduced-turbulence boundary layer at the membrane surface, which canincrease the osmotic pressure at the membrane and thus can reducepermeate flow). The concentration polarization effects can inhibit thefeed stream water from passing through the membrane as permeate, thusdecreasing the recovery ratio, e.g., the ratio of permeate to appliedfeed stream.

[0054] Reverse osmosis systems can employ a variety of different typesof actual membranes. Such commercial membrane element types include,without limitation, hollow fiber membrane elements, tubular membraneelements, spiral-wound membrane elements, plate and frame membraneelements, and the like, some of which are described in more detail in“The Nalco Water Handbook,” Second Edition, Frank N. Kemmer ed.,McGraw-Hill Book Company, New York, N.Y., 1988; see particularly Chapter15 entitled “Membrane Separation”. It should be appreciated that asingle membrane element may be used in a given membrane filtrationsystem, but a number of membrane elements can also be used depending onthe industrial application.

[0055] A typical reverse osmosis system is described as an example ofmembrane filtration and more generally membrane separation. Reverseosmosis uses mainly spiral wound elements or modules, which areconstructed by winding layers of semi-permeable membranes with feedspacers and permeate water carriers around a central perforated permeatecollection tube. Typically, the modules are sealed with tape and/orfiberglass over-wrap. The resulting construction has one channel whichcan receive an inlet flow. The inlet stream flows longitudinally alongthe membrane module and exits the other end as a concentrate stream.Within the module, water passes through the semi-porous membrane and istrapped in a permeate channel which flows to a central collection tube.From this tube it flows out of a designated channel and is collected.

[0056] In practice, membrane modules are stacked together, end to end,with inter-connectors joining the permeate tubes of the first module tothe permeate tube of the second module, and so on. These membrane modulestacks are housed in pressure vessels. Within the pressure vessel feedwater passes into the first module in the stack, which removes a portionof the water as permeate water. The concentrate stream from the firstmembrane becomes the feed stream of the second membrane and so on downthe stack. The permeate streams from all of the membranes in the stackare collected in the joined permeate tubes.

[0057] Within most reverse osmosis systems, pressure vessels arearranged in either “stages” or “passes.” In a staged membrane system,the combined concentrate streams from a bank of pressure vessels aredirected to a second bank of pressure vessels where they become the feedstream for the second stage. Commonly systems have 2 to 3 stages withsuccessively fewer pressure vessels in each stage. For example, a systemmay contain 4 pressure vessels in a first stage, the concentrate streamsof which feed 2 pressure vessels in a second stage, the concentratestreams of which in turn feed 1 pressure vessel in the third stage. Thisis designated as a “4:2:1” array. In a staged membrane configuration,the combined permeate streams from all pressure vessels in all stagesare collected and used without further membrane treatment. Multi-stagesystems are used when large volumes of purified water are required, forexample for boiler feed water. The permeate streams from the membranesystem may be further purified by ion exchange or other means.

[0058] In a multi-pass system, the permeate streams from each bank ofpressure vessels are collected and used as the feed to the subsequentbanks of pressure vessels. The concentrate streams from all pressurevessels are combined without further membrane treatment of eachindividual stream. Multi-pass systems are used when very high puritywater is required, for example in the microelectronics or pharmaceuticalindustries.

[0059] It should be clear from the above examples that the concentratestream of one stage of an RO system can be the feed stream of anotherstage. Likewise the permeate stream of a single pass of a multi-passsystem may be the feed stream of a subsequent pass. A challenge inmonitoring systems such as the reverse osmosis examples cited above isthat there are a limited number of places where sampling and monitoringcan occur, namely the feed, permeate and concentrate streams. In some,but not all, systems “inter-stage” sampling points allowsampling/monitoring of the first stage concentrate/second stage feedstream. Similar inter-pass sample points may be available on multi-passsystems as well.

[0060] In contrast to cross-flow filtration membrane separation systems,conventional filtration of suspended solids can be conducted by passinga feed fluid through a filter media or membrane in a substantiallyperpendicular direction. This effectively creates one exit stream duringthe service cycle. Periodically, the filter is backwashed by passing aclean fluid in a direction opposite to the feed, generating a backwasheffluent containing species that have been retained by the filter. Thusconventional filtration produces a feed stream, a purified stream and abackwash stream. This type of membrane separation is typically referredto as dead-end flow separation and is typically limited to theseparation of suspended particles greater than about one micron in size.

[0061] Cross-flow filtration techniques, on the other hand, can be usedfor removing smaller particles (generally about one micron in size orless), colloids and dissolved solutes. Such types of cross-flow membraneseparation systems can include, for example, reverse osmosis,nanofiltration, ultrafiltration, microfiltration, electrodialysis or thelike. Reverse osmosis can remove even low molecular weight dissolvedspecies that are at least about 0.0001 to about 0.001 microns in minimumdiameter, including, for example, ionic and nonionic species, lowmolecular weight molecules, water-soluble macromolecules or polymers,suspended solids, colloids, and such substances as bacteria and viruses.

[0062] The fluorogenic agent added to the membrane system must be amolecule or species that undergoes a change in its fluorescent signalupon interaction with a broad population of microbiological organisms.It should be appreciated by those skilled in the art that environmentalfactors, such as pH and temperature, may affect the fluorescent signaland should be accounted for and corrected accordingly. Suitablefluorogenic agents, include, but are not limited to:

[0063] acetic acid ester of pyrene 3,6,8-trisulfonic acid;

[0064] 3-carboxyumbelliferyl β-D-galactopyranoside;

[0065] 3-carboxyumbelliferyl β-D-glucuronide;

[0066] 9H-(1,3-dichloro-9,0-dimethylacridine-2-one-7-yl), D-glucuronide;

[0067] resorufin β-D-galactopyranoside;

[0068] 7-hydroxy-3H-phenoxazin-3-one 10-oxide (hereinafter “resazurin”);

[0069] 7-hydroxy-3H-phenoxazin-3-one 10-oxide, sodium salt (hereinafter“resazurin, sodium salt”);

[0070] 4-methylumbelliferyl phosphate (“4MUP”);

[0071] 4-methylumbelliferyl β-D-glucuronide;

[0072] pyranine phosphate;

[0073] pyrene 3,6,8-trisulfonic acid 1-phosphate; and

[0074] all like fluorogenic agents, derivatives and combinationsthereof.

[0075] The preferred fluorogenic agents include resazurin, 4MUP,pyranine phosphate and combinations thereof. Resazurin is the mostpreferred fluorogenic agent.

[0076] It should be appreciated by those skilled in the art that thefluorogenic agents are either commercially available (for example,resazurin, sodium salt is available from ALDRICH® of Milwaukee, Wis.) orcapable of being synthesized using procedures reported in the literature(for example, as is the case with pyranine phosphate).

[0077] The selection of fluorogenic agent and inert fluorescent tracerare made based on the membrane system being monitored and the purpose ofthe monitoring. Factors influencing the permeability of a materialthrough a membrane include the following:

[0078] a) Size of the material and size of the pores in the membrane;

[0079] b) Charge of the material and charge (or lack thereof) of themembrane;

[0080] c) Tendency of the material to adsorb on the surface of themembrane, rather than pass through the pores of the membrane;

[0081] d) Concentration Differentials between the material on one sideof the membrane and the material on the other side of the membrane; and

[0082] e) Residence times of the feed stream containing the materialbeing in contact with the membrane.

[0083] Persons of ordinary skill in the art of membranes know how to setup and run the routine tests necessary to determine whether a particularfluorogenic agent, by itself, in its reacted form or even in combinationwith a particular inert fluorescent tracer is capable of passing throughthe pores in a membrane. It is required, in order to optimally conductthe method of the first and second aspect of the instant claimedinvention that it is known, in advance, whether a fluorogenic agent iscapable of passing through the pores of the membrane. It is alsorequired, in order to optimally conduct the method of the second aspectof the instant claimed invention whether the inert fluorescent tracer iscapable of passing through the pores of the membrane.

[0084] It should be appreciated that the amount of fluorogenic agent tobe added to the membrane separation process that is effective withoutbeing grossly excessive will vary with a respect to a variety of factorsincluding, without limitation, the monitoring method selected, theextent of background interference associated with the selectedmonitoring method, the magnitude of the expected tracer(s) concentrationin the feedwater and/or concentrate, the monitoring mode (such as anon-line continuous monitoring mode), and other similar factors. In anembodiment, the amount of the fluorogenic agent added to the feed streamis an amount capable of determining microbial activity. In anembodiment, an effective amount of fluorogenic agent ranges from about 5ppt to about 500 ppm, preferably from about 0.5 ppb to about 5 ppm, andmore preferably from about 5 ppb to about 500 ppb. When the salt form ofthe agent (e.g., resazurin, sodium salt) is added to the industrialwater system, the calculation of ppm is based on the active amount ofthe fluorogenic agent present.

[0085] The cost of the fluorogenic agent also places a practical upperlimit on the amount of fluorogenic agent added to the system. Additionalfactors influencing fluorogenic agent addition to the system include thetype of fluorogenic agent, the amount of liquid continuously lost andreplenished within the membrane separation system and the type of fluidscontained within the membrane separation system.

[0086] In an embodiment, it is preferred that the fluorogenic agent ofthe present invention meet the following criteria:

[0087] 1. Not be adsorbed by the membrane in any appreciable amount;

[0088] 2. Not degrade the membrane or otherwise hinder its performanceor alter its composition;

[0089] 3. Be detectable on a continuous or semi-continuous basis andsusceptible to concentration measurements that are accurate, repeatableand capable of being performed on feedwater, concentrate water, permeatewater or other suitable media or combinations thereof;

[0090] 4. Be substantially foreign to the chemical species that arenormally present in the water of the membrane separation systems inwhich inert tracers may be used;

[0091] 5. Be substantially impervious to interference from, or biasingby, the chemical species that are normally present in the water ofmembrane separation systems in which inert tracers may be used;

[0092] 6. Be substantially impervious to any of its own potentialspecific or selective losses from the water of membrane separationsystems, including selective permeation of the membrane;

[0093] 7. Be compatible with all treatment agents employed in the waterof the membrane separation systems in which inert tracers may be used,and thus in no way reduce the efficacy thereof;

[0094] 8. Be compatible with all components of its formulation; and

[0095] 9. Be relatively nontoxic and environmentally safe, not onlywithin the environs of the water or the membrane separation system inwhich it may be used, but also upon discharge therefrom.

[0096] It should be appreciated that the present invention only requiresa single fluorogenic species to exist with the introduction of thefluorogenic agent as long as that species is reacted and can be measuredbetween the point of introduction and the point of measurement.

[0097] The fluorogenic agents suitable for use in the instant claimedprocess must have a detectable fluorescent signal prior to theirreacting with the microorganisms and also must have a differentdetectable fluorescent signal after they have reacted with themicroorganisms.

[0098] It is believed, without intending to be bound thereby, thatenzymes synthesized by the living microbiological organisms in themembrane separation can act upon the fluorogenic agents. This activitycauses a change in the fluorescent signal of the fluorogenic agent. Bymonitoring the fluorescent signal, microbiological activity duringmembrane separation can be monitored. As previously discussed, themethods of the present invention are capable of monitoringmicrobiological activity from both planktonic and sessile populations,in contrast to methods known in the art.

[0099] The fluorogenic agent can be fed either by itself or incombination with another membrane separation agent, such as an inertfluorescent tracer, a treatment agent, such as scale and corrosioninhibitors, like agents and combinations thereof. By “treatmentchemicals and/or agents” is meant without limitation treatment chemicalsthat enhance membrane separation process performance, antiscalants thatretard/prevent membrane scale deposition, antifoulants thatretard/prevent membrane fouling, biodispersants and microbial-growthinhibiting agents, such as biocides and cleaning chemicals that removemembrane deposits.

[0100] In an embodiment, the inert fluorescent tracer material is usedto determine the concentration of fluorogenic agent present and byknowing that concentration it is possible to operate the system so thata desired level of fluorogenic agent is fed accurately. See U.S. Pat.Nos. 4,783,314; 4,992,380 and 5,041,386, which are incorporated hereinby reference.

[0101] The meaning of the term “inert”, as used herein, is that an inertfluorescent tracer is not appreciably or significantly affected by anyother chemistry in the system, or by the other system parameters such asmicrobiological activity, biocide concentration and scale inhibitorconcentration. To quantify what is meant by “not appreciably orsignificantly affected”, this statement means that an inert fluorescentcompound has no more than a 10% change in its fluorescent signal, underconditions normally encountered in industrial water systems. Conditionsnormally encountered in industrial water systems are known to people ofordinary skill in the art of industrial water systems.

[0102] Inert fluorescent tracer materials suitable for use with thefluorogenic agents that are used in the instant claimed invention musthave the property of having their unique fluorescent signal bedetectably different than the fluorescent signals of the fluorogenicagent and the reacted fluorogenic agent. This means that the fluorescentsignal of the fluorogenic agent and the fluorescent signal of thereacted fluorogenic agent must both be detectably different than that ofthe inert fluorescent tracer material.

[0103] Suitable inert fluorescent tracer materials are the mono-, di-and tri-sulfonated naphthalenes, including their known water-solublesalts; and the known sulfonated derivatives of pyrene, such as1,3,6,8-pyrenetetrasulfonic acid, along with the known water-solublesalts of all of these materials, and Acid Yellow 7 (Chemical AbstractService Registry Number 2391-30-2, for 1H-Benz(de)isoquinoline-5-sulfonic acid, 6-amino-2,3-dihydro-1,3-dioxo-2-p-tolyl-,monosodium salt (8CI)).

[0104] It should be appreciated that the amount of inert fluorescenttracer to be added to the membrane separation process that is effectivewithout being grossly excessive will vary with a respect to a variety offactors including, without limitation, the monitoring method selected,the extent of background interference associated with the selectedmonitoring method, the magnitude of the expected tracer(s) concentrationin the feedwater and/or concentrate, the monitoring mode (such as anon-line continuous monitoring mode), and other similar factors. In anembodiment, the dosage of each of an inert fluorescent tracer(s) to thefeed water of the membrane separation system includes an amount that isat least sufficient to provide a measurable concentration of the inertfluorescent tracer in the concentrate at steady state of at least about5 ppt, and preferably at least about 1 part per billion (“ppb”) or about5 ppb or higher, such as, up to about 100 ppm or about 200 ppm, or evenas high as about 1000 ppm in the concentrate or other effluent stream.In an embodiment, the amount of inert fluorescent tracer ranges fromabout 5 ppt to about 1000 ppm, preferably from about 1 ppb to about 50ppm, and more preferably from about 5 ppb to about 50 ppb.

[0105] The fluorescent signals of the fluorogenic agent, the reactedfluorogenic agent and the inert fluorescent tracer material can bemeasured in a variety of different and suitable ways, including by usingcommercially available fluorometers. Fluorometers with suitable filtersmay be used to measure the fluorescent signal or signals derived fromthe fluorogenic agent, the reacted fluorogenic agent and the inertfluorescent tracer.

[0106] Examples of fluorometers that may be used in the practice of thisinvention include the TRASAR® 350 fluorometer, the TRASAR® 8000fluorometer and the Modular Fluorometer (U.S. Pat. No. 6,369,894), allavailable from Nalco; the Hitachi F-4500 fluorometer (available fromHitachi through Hitachi Instruments Inc. of San Jose, Calif.); the JOBINYVON FluoroMax-3 “SPEX” fluorometer (available from JOBIN YVON Inc. ofEdison, N.J.); and the Gilford Fluoro-IV spectrophotometer or the SFM 25(available from Bio-tech Kontron through Research InstrumentsInternational of San Diego, Calif.). It should be appreciated by thoseskilled in the art that the fluorometer list is not comprehensive and isintended only to show examples of fluorometers. Other commerciallyavailable fluorometers and modifications thereof can also be used inthis invention.

[0107] A fluorometer needs to be placed so that it can detect thefluorescent signal in whichever of the three streams that it is possibleand desired to detect the fluorescent signal of the fluorogenic agent,the reacted fluorogenic agent and the inert fluorescent tracer. Forinstance, to measure the fluorescent signal of the fluorogenic agent,the reacted fluorogenic agent and an inert fluorescent tracer, it wouldbe necessary to have three separate fluorometers set up, one to detectthe fluorescent signal of each moiety. Therefore, it would be necessaryto have three separate fluorometers set up in each of the three streams,feed stream, first stream (aka the permeate stream) and the secondstream (aka the concentrate stream).

[0108] The alternative to having three separate fluorometers is to haveone fluorometer set up that is capable of detecting the fluorescentsignals of all three moieties; such as the Modular Fluorometer describedand claimed in U.S. Pat. No. 6,369,894, issued Apr. 9, 2002.

[0109] Measuring the fluorescent signals of the fluorogenic agent, thereacted fluorogenic agent and the inert fluorescent tracer, is a knownprocedure to someone skilled in the art of fluorometry. For example, thefluorescent properties of the fluorogenic agent resazurin are well knownboth in its unreacted state and in its reacted “resorufin” state. In apreferred embodiment, the membrane separation system is sampled atlocations such that grab samples do not have to be taken formeasurements via the fluorometers. When the sampling components for thefluorometers are located within the membrane separation system, thistype of sampling is typically referred to as an in-line measurement.

[0110] An in-line measurement is one that is taken without interruptingthe flow of the system being measured. Because the sample components forthe fluorometer(s) are positioned in-line when conducting an in-linemeasurement, the sample they are monitoring accurately reflects theentire membrane separation system and, as such, the information gleanedfrom conducting this method accurately reflects both the planktonic andsessile microbiological organism populations. In-line measurementovercomes the problems associated with grab sampling and the need toremove a sample from the aqueous stream for later testing. Also, thereacted and unreacted forms of the fluorogenic agents are tested on areal-time basis, wherein an almost instantaneous fluorescence reading ofthe two fluorogenic agents will provide an indication of microbialactivity.

[0111] Notwithstanding the fact that in-line measurement is thepreferred way of conducting the method of the instant claimed invention,it is possible to conduct the method of the instant claimed inventionusing a grab sampling technique suitable to secure samples of theindustrial water system. If a grab sampling technique is used, amechanism should necessarily be provided to convey the grab sample tothe fluorometer in a reasonable length of time such that the datareceived from the fluorometer accurately reflects the current microbialgrowth status of the membrane separation system.

[0112] In an embodiment, a fluorescent signal ratio is used as anindication of microbial activity and thus an indication of biofoulingduring membrane separation. By calculating the ratio as opposed tosimply measuring an absolute value of fluorescent signals, informationis obtained that is independent of fluorogenic agent concentration. Inanother embodiment, the ratio can increase the sensitivity due to thefact that the microbiological organisms convert fluorogenic reagent toreacted fluorogenic agent with the ratio increase being due to both thedecrease in the fluorescent signal of the unreacted fluorogenic agentand increase in the fluorescent signal of the reacted fluorogenic agent(e.g., the product).

[0113] The ratio of the fluorescent signal of the fluorogenic agent tothe fluorescent signal of the reacted fluorogenic agent is:${ratio} = \frac{{fluorescent}\quad {signal}\quad {of}\quad {reacted}\quad {fluorogenic}\quad {agent}}{{fluorescent}\quad {signal}\quad {of}\quad {fluorogenic}\quad {agent}}$

[0114] The ratio is a unitless number. The ratio can be calculatedmanually or with a calculator or with a computer program. For ease ofuse, it is preferable that the ratio be calculated using an appropriatecomputer program such that a record of the ratio can be continuouslycalculated at set intervals. The rate of change of the ratio can then beused to determine the rate of increase of biofouling.

[0115] If necessary, other ratios can be determined and used toadvantage. For example, the ratio of the fluorescent signal of thereacted fluorogenic reagent to the fluorescent signal of the inerttracer (second aspect of the instant claimed invention) could also beused to glean valuable information about the amount of biofoulingoccurring in the membrane separation system.

[0116] The present invention can include a controller to monitor and/orcontrol the operating condition or performance of the membraneseparation system based on the measurable fluorescent signals of thefluorogenic agent, reacted fluorogenic agent, and the inert fluorescenttracer. In this regard, biofouling is monitored in the membraneseparation system by measuring the change in the signal of thefluorogenic agent or the reacted fluorogenic agent with respect to thesignal of the inert fluorescent tracer and then comparing the resultantratio to the corresponding ratio in the feed stream or the correspondingratio at time zero, i.e., right after the membrane has been cleaned. Thecontroller can be configured and/or adjusted in a variety of differentand suitable ways.

[0117] For example, the controller can be in contact with thefluorometer to process the detection signal (e.g., filter noise from thesignal) in order to enhance the detection of the fluorescent signal(s)derived from the fluorogenic agent, reacted fluorogenic agent and/or theinert fluorescent tracer. Further, the controller can be adjusted tocommunicate with other components of the membrane separation system. Thecommunication can be either hard wired (e.g., electrical communicationcable), a wireless communication (e.g., wireless RF interface), apneumatic interface or the like.

[0118] In this regard, the controller can be used to control theperformance of membrane separation by determining the optimal amount ofbiocontrol treatment needed. “Biochemical treatment” includes biocides,biocontrol agents, biocontrol methods and combinations thereof. Forexample, the controller can communicate with a feed device (not shown)in order to control various biofouling control chemicals or biofoulingcontrol devices or process parameters. In an embodiment, the controlleris capable of adjusting the feed rate of biocontrol agent(s) added tothe feed stream during membrane separation based on the fluorescence ofthe fluorogenic agent and reacted fluorogenic agent that are measured.In another embodiment, the addition of biocontrol agents are controlledbased on the determination of the ratio of reacted fluorogenic agent tofluorogenic agent. In another embodiment, the addition of biocides arecontrolled based on the determination of the rate of change of ratio ofreacted fluorogenic agent to fluorogenic agent.

[0119] A real-time determination of the fluorescence of the fluorogenicagent(s) enables immediate evaluation of the microbial activity as wellas the efficiency of the current biocontrol agent dosage and/or the needfor an additional feed of the biocontrol agent. Real-time determinationof biological activity enables the process to add biocontrol agents onan as needed basis. Adding excess biocontrol agents, greater than whatis needed to control the microbial activity is avoided when thebiofouling condition is accessed on a real-time basis. Thus, the use ofbiocontrol agents can be administered at determined effective levels,which results in the correct amount being used. Additionally, theeffectiveness of a biocontrol agent feed can be evaluated on a real-timebasis and the dosage can be increased or decreased depending upon thereal-time reading.

[0120] When the method of the instant claimed invention is conducted inthe presence of biocontrol agents, certain adjustments have to be made.People of ordinary skill in the art know what biocides are used inmembrane separation systems. In an embodiment, biocides added inresponse to unacceptable levels of microbial activity include oxidizingbiocides, non-oxidizing biocides and combinations thereof.

[0121] It is extremely uncommon for oxidizing biocides to be used inreverse osmosis membrane separation systems or in other membrane systems(for example, polyamide membranes) that are not oxidant tolerant.However, when used with membranes that are oxidant tolerant, oxidizingbiocides can be used. Persons of ordinary skill in the art of membraneswould be aware of which combination of biocide and membrane material ofconstruction would be acceptable.

[0122] When oxidizing biocides are selected, suitable oxidizing biocidesinclude, but are not limited to:

[0123] BCDMH (92.5%, 93.5%, 98%), which is either1,3-dichloro-5,5-dimethylhydantoin and 1-bromo-3-chloro-5,5-dimethylhydantoin (CAS Registry #16079-88-2) or a mixture thereof;

[0124] bleaches, including stabilized bleaches;

[0125] bromine, including stabilized bromine;

[0126] calcium hypochlorite (CAS Registry #7778-54-3) “Cal Hypo” (68%);

[0127] chlorine, including stabilized chlorine (8.34%);

[0128] H₂O₂/PAA (21.7%/5. 1%) which is hydrogen peroxide (CAS Registry#7722-84-1)/peracetic acid (CAS Registry #79-21-0);

[0129] hypobromite;

[0130] hypobromous acid;

[0131] iodine;

[0132] organobromines;

[0133] NaBr (42.8%, 43%, 46%) which is sodium bromide;

[0134] NaOCl (10%, 12.5%) which is sodium hypochlorite (CAS Registry#7681-52-9);

[0135] and mixtures thereof.

[0136] Suitable non-oxidizing biocides include, but are not limited to,

[0137] ADBAC Quat (10%, 40%(CAS Registry #68391-0-5), 80%)—alkyldimethyl benzyl ammonium chloride, also known as “quat”;

[0138] ADBAC quat(15%)/TBTO (tributyl tin oxide 5%);

[0139] ADBAC(12.5%)/TBTO (2.5%), (ADBAC Quat/bis tributyl tin oxide)(CAS Registry #56-35-9);

[0140] carbamates (30%), of formula T₂NCO₂H, where T₂ is a C₁-C₁₀ alkylgroup;

[0141] copper sulfate (80%);

[0142] DBNPA (20%, 40%), which is 2,2-dibromo-3-nitrilopropionamide (CASRegistry #10222-01-2);

[0143] DDAC Quat (50%) which is didecyl dimethyl ammonium chloride quat;

[0144] DPEEDBAC Quat (1%) which is(2-(2-p-(diisobutyl)phenoxy)ethoxy)ethyl dimethyl, dimethyl benzyl;

[0145] glutaraldehyde (15%, 45%), CAS Registry #111-30-8;

[0146] glutaraldehyde (14%)/ADBAC quat (2.5%);

[0147] HHTHT—hexahydro-1,3,5-tris (2-hydroxyethyl)-5-triazine (78.5%);

[0148] isothiazolones (1.5%, 5.6%)—a mixture of5-chloro-2-methyl-4-isothiazoline-3-one (CAS Registry #26172-55-4) and2-methyl;-4-isothiazoline-3-one (CAS Registry #2682-20-4);

[0149] MBT (10%)—methylene bis thiocyanate;

[0150] polyquat (20%, 60%), a polymeric quaternary compound; polyaminesand salts thereof—polymeric amine compounds;

[0151] terbutylazine (4%,44.7%)—2-(tert-butylamino)-4-chloro-6-ethylamino-5-triazine (CASRegistry #5915-41-3);

[0152] TMTT (24%)—tetramethylthiuram disulfide;

[0153] and mixtures thereof.

[0154] Other types of biocontrol agents include bio-dispersants,bio-detergents, chaotropic agents, surfactants, chelating agents,enzymatic cleaners, and other chemicals that kill bacteria or interferewith bacterial and EPS attachment and bacteria colonization processes.

[0155] Biocontrol methods, such as mechanical means to disrupt biofilmintegrity including ultrasound, electric fields, air backwashes, etc.can also be used.

[0156] It has been found that all of the fluorogenic reagents suitablefor use in the instant claimed invention are susceptible to degradationof varying degrees in the presence of oxidizing biocides. When themethod of the instant claimed invention is used in an industrial watersystem where these oxidizing biocides are present it is important to addthe fluorogenic reagent to the industrial water system at a point thatis as far as possible away from the point where the oxidizing biocide isadded to the industrial water system. Even when the fluorogenic reagentand the oxidizing biocide are added to the industrial water system atpoints as far apart as possible it is known that the oxidizing biocidewill quench the fluorescent signal of both the fluorogenic reagent andthe reacted fluorogenic reagent. The quenched fluorescent signals cannotaccurately reflect the current status of the microbiological activity inthe industrial water system.

[0157] Because membrane systems have very short holding times, theinterference of biocontrol agents, specifically oxidizing biocidebiocontrol agents with the fluorogenic agent is not expected to pose anyproblems in most practical applications. This will be true as long asthe biocontrol agent is not applied during the introduction of thefluorogenic agent. In practice, the biocontrol agent may be addedintermittently, and thus avoid direct contact with the fluorogenicagent.

[0158] Although membrane separation systems are often employed for thepurification of water, or the processing of aqueous streams, the systemsof the present invention are not limited to the use of an aqueousinfluent. In an embodiment, the influent may be another fluid, or acombination of water and another fluid. The operational principles ofmembrane separation systems and processes of the present invention arenot so governed by the nature of the influent that the present inventioncould not be employed with influents otherwise suitable for waterpurification in a given membrane separation system. The descriptions ofthe invention above that refer to aqueous systems are applicable also tononaqueous and mixed aqueous/nonaqueous systems.

[0159] The foregoing descriptions of the present invention at timesrefer specifically to aqueous influents and effluents, and the use of anaqueous system for describing a membrane filtration system and theoperation of the present invention therein is exemplitive. A person ofordinary skill in the art, given the disclosures of the presentspecification, would be aware of how to apply the foregoing descriptionsto nonaqueous membrane filtration systems.

[0160] It should be appreciated that the present invention is applicableto all industries that can employ membrane separation processes. Forexample, the different types of industrial water systems in which themethod of the present invention can be applied generally include rawwater processes, waste water processes, industrial water processes,municipal water treatment, food and beverage processes, pharmaceuticalprocesses, electronic manufacturing, utility operations, pulp and paperprocesses, mining and mineral processes, transportation-relatedprocesses, textile processes, plating and metal working processes,laundry and cleaning processes, leather and tanning processes, and paintprocesses.

[0161] In particular, food and beverage processes can include, forexample, dairy processes relating to the production of cream, low-fatmilk, cheese, specialty milk products, protein isolates, lactosemanufacture, whey, casein, fat separation, and brine recovery fromsalting cheese. Uses relating to the beverage industry including, forexample, fruit juice clarification, concentration or deacidification,alcoholic beverage clarification, alcohol removal for low-alcoholcontent beverages, process water; and uses relating to sugar refining,vegetable protein processing, vegetable oil production/processing, wetmilling of grain, animal processing (e.g., red meat, eggs, gelatin, fishand poultry), reclamation of wash waters, food processing waste and thelike.

[0162] Examples of industrial water uses as applied to the presentinvention include, for example, boiler water production, process waterpurification and recycle/reuse, softening of raw water, treatment ofcooling water blow-down, reclamation of water from papermakingprocesses, desalination of sea and brackish water for industrial andmunicipal use, drinking/raw/surface water purification including, forexample, the use of membranes to exclude harmful micro-organisms fromdrinking water, polishing of softened water, membrane bio-reactors,mining and mineral process waters.

[0163] Examples of waste water treatment applications with respect tothe inert tracer monitoring methods of the present invention include,for example, industrial waste water treatment, biological wastetreatment systems, removal of heavy metal contaminants, polishing oftertiary effluent water, oily waste waters, transportation relatedprocesses (e.g., tank car wash water), textile waste (e.g., reagent,adhesives, size, oils for wool scouring, fabric finishing oils), platingand metal working waste, laundries, printing, leather and tanning, pulpand paper (e.g., color removal, concentration of dilute spent sulfiteliquor, lignon recovery, recovery of paper coatings), chemicals (e.g.,emulsions, latex, pigments, paints, chemical reaction by-products), andmunicipal waste water treatment (e.g., sewage, industrial waste).

[0164] Other examples of industrial applications of the presentinvention include, for example, semiconductor rinse water processes,production of water for injection, pharmaceutical water including waterused in enzyme production/recovery and product formulation, andelectro-coat paint processing. The present invention provides methodsfor monitoring and/or controlling biofouling in membrane separationsystems that are capable of removing solutes and/or other impuritiesfrom feed streams, such as aqueous feed streams, which are suitable foruse in a number of different industrial applications. More specifically,the methods of the present invention can monitor and/or controlbiofouling in membrane separation systems based on monitoring theactivity of a fluorogenic agent(s) which has been added to the feedstream. In this regard, biofouling due to microbial growth duringmembrane separation can be evaluated with a high degree of selectivity,specificity and accuracy such that the performance of the membraneseparation system can be effectively optimized.

[0165] Membrane separation systems and the monitoring thereof are uniquebecause of the following considerations.

[0166] 1. Systems are constructed with limited flexibility in terms ofwhere monitoring may be done and/or where samples may be collected.Moreover, different types of fouling are usually located in specificsegments of the membrane. For example, biofouling is usually predominantat the front end of the membrane system.

[0167] 2. Membrane separation systems include a concentrationpolarization layer that forms as water is permeated through the barrier.This is not present in other water treatment systems, such as coolingwater systems.

[0168] 3. Because it is essential that the surface of the membraneremain clean, a relatively small amount of biofouling or fineprecipitate can cause a significant performance loss. The performanceloss in a membrane is, thus, more sensitive as compared to cooling watertreatment. In this regard, performance loss in a membrane can occur at afilm thickness appreciably lower than that required for heat transferloss to occur in a cooling water system. Moreover, due to the small flowchannels, the fouling, when not controlled or removed earlier, willcontinue to accelerate and it will be more difficult, if not impossible,to clean the membrane back to its original performance capability. Ashortened membrane life is typically caused by this delay in detectionof fouling.

[0169] 4. The thin, semi-permeable films (polymeric, organic orinorganic) are sensitive to degradation by chemical species, includingthe chemicals secreted from the cell growth process and the cleaningchemicals. Products which contact membrane surfaces must be compatiblewith the membrane chemistry to avoid damaging the surface and therebydegrading performance.

[0170] 5. Chemical treatments used in membrane systems must bedemonstrated to be compatible with the membrane material prior to use.Damage from incompatible chemicals can result in immediate loss ofperformance and perhaps degradation of the membrane surface. Suchimmediate, irreversible damage from a chemical treatment is highlyuncommon in cooling water systems.

[0171] 6. In membrane separation systems, there is a continuous feedstream and at least one continuous discharge stream. The holding time ofa membrane separation system is very short, i.e., on the order of a fewseconds to a few minutes.

EXAMPLES

[0172] The following examples are intended to be illustrative of thepresent invention and to teach one of ordinary skill how to make and usethe invention. These examples are not intended to limit the invention orits protection in any way.

Example 1

[0173] A test membrane system was used to demonstrate the monitoring ofbiofilm growth on a reverse osmosis membrane. The system was a test unit(SEPA CF membrane cell) manufactured by Osmonics. The unit included anew (i.e. clean) flat sheet of membrane measuring 3.5 inches×5 inches.Feed water at 360 psig was forced into the feed channel of the membranehousing at about 150 milliliters per minute (ml/min). As the feed watertraveled across the membrane towards the reject exit channel, pressureforced the water molecules through the membrane into the permeatechannel at about 15 ml/min. Exit water on the reject channel was about360 psig. The pressure drop reported was the pressure at the feedchannel minus the pressure at the reject channel.

[0174] The permeate flow as percentage of feed flow was 10%, which istypical for a reverse osmosis membrane element. The feed water had aconductivity of 3.9 milli-Siemens/cm. The permeate water had aconductivity of 0.2 milli-Siemens/cm. The operating temperature was 78°F. and the pH was 7.

[0175] In the test membrane system, a seeding of bacteria was performedand, thus, the microbial biofouling began almost immediately afterseeding. The experiment was run for a period of about 90 to 170 hoursafter seeding. Variables affected by fouling (e.g., permeate flow,pressure differential between feed channel and reject channel, andpermeate conductivity) were measured. The surface microbial enumerationof the membrane was performed at the end of the run.

[0176] To demonstrate the use of the fluorogenic agent, a solution ofabout 2 ppm (parts per million) resazurin was fed at 3 ml/min into thefeed stream (which had a flow rate of about 150 ml/min) before itentered the membrane feed channel for a period of about five minutes.The concentration of the resazurin after mixing with the feed water wasabout 40 ppb (parts per billion).

[0177] The fluorescence measurement was conducted in the reject streamwith an on-line fluorometer, Nalco TRASAR® 350, and with a bench-topfluorometer, Jobin Yvon Spectrometer Fluoro-MAX 3. The residence time ofthe resazurin chemical in the membrane system was on the order of aboutone minute. The fluorescence measurement reached a steady stategenerally in about three minutes after commencement of the resazurinfeed. The 5-minute feeding and measurement of the fluorogenic agent wasrepeated periodically during the course of the experiment. TABLE 1Microbial Permeate enumeration TRASAR Permeate Pressure conductivity onTime 350 flow drop (milli- membrane* (Hr) (Ratio) (ml/min) (inch water)Siemen/cm) (cfu/cm²) 1 1.0 15 0.8 0.17 — 28 1.4 13 0.8 0.13 — 54 1.9 100.9 0.16 — 72 2.0 7 0.9 0.20 — 144 2.3 3 1.7 0.29 4 × 10⁸

[0178] TABLE 2 TRASAR 350 Fluoro-MAX 3 Time 583 nm 634 nm 583 nm 634 nmHr Ratio Rf Rz Ratio Rf Rz 1 1.0 39 41 1.0 24300 23900 28 1.4 70 50 1.544100 29600 54 1.9 96 51 2.0 59600 29500 72 2.0 110 55 2.2 71600 33100144 2.3 133 58 2.4 73200 30900

[0179] As shown above in Tables 1 and 2, the Ratio of the fluorescenceof the reacted fluorogenic agent (e.g., resorufin (Rf) to thefluorescence of the fluorogenic agent (e.g., resazurin (Rz)) increasedwith microbial growth and accumulation during the course of theexperiment. It should be appreciated that the values for Rf and Rz inTables 1 and 2 represent an intensity of the fluorescence signal asmeasured which corresponds to an amount of the reacted fluorogenic agentand the fluorogenic agent. Furthermore, Tables 1 and 2 demonstrate theexistence of a correlation between the Ratio and the membrane separationperformance variables measured by permeate flow, pressure drop andpermeate conductivity.

Example 2

[0180] A batch test was performed to quantify the reaction betweenmicrobial activity and resazurin, a fluorogenic agent. The test wasconducted in a minimal medium solution containing 100 ppm glycerol, 7grams/liter (g/L) K₂HPO₄, 0.1 g/L MgSO₄.7H₂O, 1 g/L (NH₄)₂.SO₄ and 0.5g/L sodium citrate. A field deposit from a reverse osmosis membrane unitwas inoculated in the medium and allowed to grow. The solution was thenincubated at 30° C. in a flask shaker at a speed of about 200revolutions per minute (rpm). Aliquots of sample were collected using asterile pipette at different time intervals. Samples collected from eachtime point were analyzed for optical density, microbial enumeration (at8, 12, and 24 hr) and allowed to react with resazurin. Optical densitywas performed with absorbance at 600 nm using a CintraSpectrophotometer. Microbial enumeration was performed with spread plateon TGE agar after eight serial dilutions of 1:10 with a sterilephosphate buffer solution. In resazurin reaction tests, samples weredosed with 50 ppb of resazurin. Aliquots of a 3.5 ml sample wereretrieved at reaction times of 1, 2, 3, 5, 10 minutes and then filteredthrough a 0.22 micrometer sterile syringe filter to stop the reaction.These reacted samples were subsequently measured for fluorescence ofresazurin and resorufin. Response of resazurin was judged by thefluorescence ratio of resorufin to resazurin. Fluorescence measurementswere conducted at excitation wavelength 550 nm and emission wavelengths583 nm and 634 nm, using a Horiba Jobin Yvon fluorometer (modelFluoroMax-3) with slit width 2.5 nm (excitation) and 2.5 nm (emission).resazurin had a peak fluorescence at 634 nm and resorufin at 583 nm.TABLE 3 Microbial Growth Optical with Fluorescence ratio of resorufin toresazurin at density Viable plate Time each reaction time for eachsample (Intensity) (absorbance) count (hour) t = 1 min t = 2 min t = 3min t = 5 min t = 10 min 600 nm cfu/ml 0 1.00 0.93 0.94 0.89 1.00 0.00014 1.29 1.42 0.95 0.94 1.31 6 1.03 1.27 0.96 1.20 0.97 0.0020 8 1.17 1.141.17 1.18 1.29 0.0094 3.10E+06 10 1.25 1.20 1.30 1.33 1.29 0.0162 121.55 1.52 1.61 1.58 1.79 0.0629 3.90E+07 24 2.81 3.11 3.22 3.50 3.750.4261 2.40E+09

[0181] As demonstrated in Table 3, the fluorescence ratio of resorufinto resazurin increased as microbial growth increased as indicated byoptical density (absorbance 600 nm) and viable cell count. The ratioalso increased as reaction time between the sample and resazurinincreased, especially when microbial density was about 10⁶cfu/ml orabove. The results show that resazurin is responsive to the microbialgrowth of a membrane biofilm.

Example 3

[0182] A batch test was performed to quantify the relationship between aresazurin dose concentration and its response to different levels ofmicrobial density during microbial growth. The test was conducted in aminimal medium solution inoculated with field reverse osmosis (RO)membrane deposit as described in Example 2. The solution was incubatedat 30° C. in a flask shaker at a speed of 200 rpm. Aliquots of samplewere collected using a sterile pipette at different time intervals.Samples collected from each time point were analyzed for opticaldensity, microbial enumeration (except time zero and 2 hours) andallowed to react with resazurin. Optical density and microbialenumeration were performed using the same methods described in Example2. In the resazurin reaction test, samples were dosed with 50 ppb, 500ppb and 5000 ppb of resazurin. Aliquots of 3.5 ml samples were retrievedafter 1 minute reaction. These reacted samples were subsequentlymeasured for fluorescence of resazurin and resorufin. The response ofresazurin was evaluated by the fluorescence ratio of resorufin toresazurin. The method of fluorescence measurements is described above inExample 2. TABLE 4 Microbial Fluorescence ratio of Relative changegrowth resorufin to resazurin at of ratio (Rf/Rzt − Optical densityViable plate time each dose concentration Rf/Rzo) (Absorbance) count(hr) 0.05 ppm 0.5 ppm 5 ppm 50 ppb OD600 cfu/ml 0 0.878 0.761 0.4000.0013 2 1.107 0.766 0.413 0.229 0.0013 4 1.241 0.927 0.445 0.363 0.00131.40E+05 6 1.217 0.892 0.456 0.338 0.0112 2.10E+05 8 1.147 0.928 0.4560.269 0.0029 4.70E+05 10 1.244 0.930 0.476 0.366 0.0052 2.30E+06 141.346 0.921 0.433 0.468 0.0223 1.90E+07 25 2.560 1.520 0.676 1.6820.2185 2.50E+09

[0183] TABLE 5 Fluorescence ratio of Rf/Rz at different microbial celldensity Log Resazurin Rz dose 2.00E+06 2.00E+07 3.00E+09 dose (ppb)(ppb) (cfu/ml) 1.70 50 1.24 1.35 2.56 2.70 500 0.93 0.92 1.52 3.70 50000.48 0.43 0.68 Linear regression Slope −0.38 −0.46 −0.94 Intercept 1.922.13 4.13 RSQ 0.9890449 0.99844 0.996395936

[0184] As indicated in Tables 4 and 5, the fluorescence ratio ofresorufin to resazurin increased as the resazurin dose decreased at eachmicrobial cell density. Furthermore, the dose of resazurin concentrationhad a logarithm linear relationship with the fluorescence ratio responsein the sample. The change of the ratio also indicated 50 ppb ofresazurin was more sensitive to microbial growth than 500 ppb and 5000ppb of resazurin. The fluorescence ratio of resorufin to resazurin wasan order of magnitude more sensitive than the optical densitymeasurement.

Example 4

[0185] This test evaluated the correlation between fluorescence responseof resazurin and biofilm (sessile cells) on a reverse osmosis (RO)membrane. The test was performed in a 350 ml fed-batch flow cell system.The system was fed with a minimal medium and inoculated with a field ROdeposit as described in Example 2. The flow cell was operated at ahydraulic residence time of about 14 hr. A recirculation pump providedmixing to the system at 195 ml/min. RO membrane coupons of 3 inch×1 inchwere submerged in the flow cell.

[0186] Membrane samples were taken periodically for sessile microbialenumeration and resazurin response test. A membrane sample was soaked ina sterile phosphate buffer solution (PBS) in a 45 ml test tube for a 5minute sonication. An aliquot of bulk from the sonicated sample was thendiluted with sterile PBS in eight serial dilutions of 1:10 before platedon TGE agar plate. The viable plate count represented sessile celldensity from the RO membrane. Resazurin of 40 ppb was dosed into thesonicated membrane sample in PBS for one minute. An aliquot of samplewas then taken to check for fluorescence at the same conditions asdescribed in Example 2. The measured fluorescence ratio of resorufin toresazurin from the sonicated membrane sample represented response tomicrobial flocs. Another membrane sample was retrieved and soaked in 45ml PBS in a test tube. The sample was dosed with 40 ppb resazurin forone minute. An aliquot of sample was then collected to measurefluorescence at the same conditions as described in Example 2. Thefluorescence ratio associated with this sample represented response tointact biofilms on the RO membrane. TABLE 6 Sessile cell density onRatio of Elapsed time RO membrane resorufin to (hr) Log (cfu/cm²)resazurin 1 3.76 0.824 24 7.41 48 7.43 0.888 72 7.73 0.888 96 9.09 0.992168 9.11 0.955 174 8.37 1.051

[0187] TABLE 7 Sessile cell Ratio of density on resorufin to RO membraneresazurin Log (cfu/cm2) biofilm flocs 7.43 0.888 0.993 7.73 0.888 1.1159.09 0.992 1.031

[0188] Table 6 illustrates that response of resazurin increases assessile cell density from RO membrane increases. Table 7 furtherdemonstrates that the response of resazurin is greater in flocs thanintact biofilms.

[0189] While the present invention is described above in connection withpreferred or illustrative embodiments, these embodiments are notintended to be exhaustive or limiting of the invention. Rather, theinvention is intended to cover all alternatives, modifications andequivalents included within its spirit and scope, as defined by theappended claims.

1. A method of monitoring biofouling in a membrane separation systemincluding a membrane capable of separating a feed stream into at least afirst stream, known as the permeate, and a second stream, known as theconcentrate, comprising the steps of: (a) selecting a fluorogenic agentwherein the selection is made such that it is known in advance whethersaid fluorogenic agent is (i) capable of traveling through the membraneinto the permeate stream, or (ii) not capable of passing through themembrane into the permeate stream; (b) adding the fluorogenic agent tothe feed stream; (c) providing one or more fluorometers to detect thefluorescent signal of the fluorogenic agent in at least one of the feedstream, the concentrate and optionally the permeate; (d) allowing thefluorogenic agent to react with at least one microorganism within themembrane separation system to form a reacted fluorogenic agent; (e)using said one or more fluorometers to detect the fluorescent signal ofat least one of the fluorogenic agent and the reacted fluorogenic agentin at least one of the feed stream and the concentrate and optionallythe permeate; and (f) using the fluorescent signal of at least one ofthe fluorogenic agent and the reacted fluorogenic agent to monitorbiofouling in the membrane separation system based on the change in thefluorescent signal of the fluorogenic agent, or the reacted fluorogenicagent or a combination of both fluorescent signals.
 2. The method ofclaim 1 wherein the membrane separation system is selected from thegroup consisting of a cross-flow membrane separation system and adead-end flow membrane separation system.
 3. The method of claim 1wherein the membrane separation system is selected from the groupconsisting of reverse osmosis, nanofiltration, ultrafiltration,microfiltration, electrodialysis, electrodeionization, pervaporation,membrane extraction, membrane distillation, membrane stripping, membraneaeration and combinations thereof.
 4. The method of claim 1 wherein thefluorogenic agent is selected from the group consisting of: acetic acidester of pyrene 3,6,8-trisulfonic acid; 3-carboxyumbelliferylβ-D-galactopyranoside; 3-carboxyumbelliferyl β-D-glucuronide;9H-(1,3-dichloro-9,0-dimethylacridine-2-one-7-yl), D-glucuronide;resorufin β-D-galactopyranoside; resazurin; resazurin, sodium salt;4-methylumbelliferyl phosphate; and combinations thereof.
 5. The methodof claim 1 wherein the fluorogenic agent is resazurin.
 6. The method ofclaim 1 wherein the fluorogenic agent is added into the feed stream inan amount from about 5 ppt to 500 ppm.
 7. The method of claim 1 whereinbiofouling is monitored by determining a ratio of the fluorescent signalof the reacted fluorogenic agent to the fluorescent signal of thefluorogenic agent in at least one of the permeate and concentrate. 8.The method of claim 1 further comprising the step of: (g) determiningthe optimal amount of biocontrol treatment based on the change in thesignal of the fluorogenic agent, or the reacted fluorogenic agent, or acombination of both signals measured; and (h) applying the optimalamount of biocontrol treatment to the membrane separation system.
 9. Themethod of claim 8 wherein the biocontrol treatment is selected from thegroup consisting of biocides, biocontrol agents, biocontrol methods andcombinations thereof.
 10. The method of claim 8 wherein the biocides areselected from the group consisting of oxidizing biocides, non-oxidizingbiocides and combinations thereof.
 11. The method of claim 8 wherein thebiocontrol treatment is selected from the group consisting ofbio-dispersants, bio-detergents, chaotropic agents, surfactants,chelating agents, enzymatic cleaners and combinations thereof.
 12. Themethod of claim 8 wherein the biocontrol treatment is selected from thegroup consisting of ultrasound, electric fields and air backwashes. 13.The method of claim 1 wherein the microorganisms are selected from thegroup consisting of planktonic microorganisms, sessile microorganismsand combinations thereof.
 14. The method of claim 1 in which the feedstream is aqueous.
 15. The method of claim 1 in which the feed stream isnon-aqueous.
 16. A method of monitoring biofouling in a membraneseparation system including a membrane capable of separating a feedstream into at least a first stream, known as the permeate, and a secondstream, known as the concentrate, comprising the steps of: (a) selectinga fluorogenic agent wherein the selection is made such that it is knownin advance whether said fluorogenic agent is (i) capable of travelingthrough the membrane into the permeate stream, or (ii) not capable ofpassing through the membrane into the permeate stream; (b) selecting aninert fluorescent tracer wherein the selection is made such that it isknown in advance whether said inert fluorescent tracer is (i) capable oftraveling through the membrane into the permeate stream, or (ii) notcapable of passing through the membrane into the permeate stream; (c)adding the fluorogenic agent and the inert fluorescent tracer to thefeed stream, wherein said fluorogenic agent and said inert fluorescenttracer are added in a known proportion to each other; (d) providing oneor more fluorometers to detect the fluorescent signal of the fluorogenicagent and the fluorescent signal of the inert fluorescent tracer in atleast one of the feed stream or the concentrate or optionally thepermeate; (e) allowing the fluorogenic agent to react with at least onemicroorganism within the membrane separation system to form a reactedfluorogenic agent; (f) using said one or more fluorometers to detect thefluorescent signal of at least one of the fluorogenic agent and thereacted fluorogenic agent and the inert fluorescent tracer in at leastone of the feed stream and the concentrate and optionally the permeate;and (g) using the fluorescent signal of at least one of the fluorogenicagent and the reacted fluorogenic agent and the inert fluorescent tracerto monitor biofouling in the membrane separation system based on thechange in the fluorescent signal of the fluorogenic agent, or thereacted fluorogenic agent or a combination of both fluorescent signalsrelative to the fluorescent signal of the inert fluorescent tracer. 17.The method of claim 16 in which said inert tracer is selected from thegroup consisting of the mono-, di- and tri-sulfonated naphthalenes,including their known water-soluble salts; the sulfonated derivatives ofpyrene, including 1,3,6,8-pyrenetetrasulfonic acid, along with the knownwater-soluble salts of all of these materials and 1H-Benz(de)isoquinoline-5-sulfonic acid, 6-amino-2,3-dihydro-1,3-dioxo-2-p-tolyl-,monosodium salt (8CI)).
 18. The method of claim 16 wherein thefluorogenic agent is selected from the group consisting of: acetic acidester of pyrene 3,6,8-trisulfonic acid; 3-carboxyumbelliferylβ-D-galactopyranoside; 3-carboxyumbelliferyl β-D-glucuronide;9H-(1,3-dichloro-9,0-dimethylacridine-2-one-7-yl), D-glucuronide;resorufin β-D-galactopyranoside; resazurin; resazurin, sodium salt;4-methylumbelliferyl phosphate; and combinations thereof.
 19. The methodof claim 16 wherein the fluorogenic agent is resazurin.
 20. The methodof claim 16 further comprising the steps of: (h) determining the optimalamount of biocontrol treatment based on the change in the signal of thefluorogenic agent, or the reacted fluorogenic agent, or a combination ofboth signals measured; and (i) applying the optimal amount of biocontroltreatment to the membrane separation system.